Project Summary/Abstract Children and young adults who were exposed to alcohol during fetal life often show fetal alcohol spectrum disorders (FASD) consisting of learning disabilities, emotional disturbances, stress abnormalities and immune incompetence. Furthermore, some grandparents who had alcohol-related medical problems often had grandchildren with a higher rate of fetal alcohol syndrome than those grandparents without alcohol-related medical problems. Epidemiological studies also have shown association of paternal alcohol consumption with deficiencies in offspring?effects which are typically found with maternal alcohol exposure. However, the process underlying the heritability of alcohol-induced health issues is not clearly understood. Our recent studies identified an epigenetic mechanism in the heritable effect of fetal alcohol on stress response abnormalities and immune dysfunctions. These studies showed that the proopiomelanocortin (Pomc) gene, which negatively controls the functions of the stress axis and stimulates the functions of the immune systems, is hypermethylated while the response of stress axis hormones and the production of the cytokine interferon-? (IFN-?) is dysregulated in fetal alcohol exposed offspring for three generations via the male germline. However, the process by which these epigenetic variants induced by fetal alcohol are transmitted from parents to offspring through multiple generations has not been established. We hypothesize that a transcription factor like sex-determining region Y (SRY), which has significant control over Pomc gene expression, undergoes heritable epigenetic modifications by fetal alcohol exposures that in turn modify the expression of the Pomc gene and its endophenotypes. The objective of this proposal is to determine the heritable epigenetic effects of alcohol exposure during the prenatal period on SRY-mediated Pomc gene expression and its endophenotypes, i.e., the stress hormone response and IFN-? production to an immune challenge, in isogenic Fischer 344 rats. This research aim will be achieved by determining the relationship between the fetal alcohol epigenetic marks on Pomc and its endophenotypes with the epigenetic marks on the Sry gene in the isogenic rat population in three generations of offspring, studying the interaction between SRY and Pomc at the molecular level, and evaluating whether increased methylation affects SRY interaction with the Pomc promoter and contributes to fetal alcohol effects on Pomc and its endophenotypes. Various state of the art techniques will be employed involving promoter methylation, promoter demethylation, promoter activity, mutational analysis, gene knock down, in vivo electroporation for gene overexpression, and stress and immune response studies. These studies will establish the process by which developmental alcohol exposure induces gene expression variants in the offspring and how they are transmitted to next generations. Knowledge on the process involved in the heritability of alcohol-induced epigenetic variants may help in establishing new biomarkers and novel epigenetic-based therapies for many alcohol-related disorders.